Although neurodegenerative disorders are prevalent in the aging human population, the molecular mechanisms underlying these diseases are not well understood. As in humans, genetic lesions have been associated with neurodegeneration in mice. Several mouse mutations exist that result in the abnormal death of embryonic or early postnatal neurons. In contrast, our studies of mice homozygous for the spontaneous mutation harlequin (Hq) have shown that this mutation causes progressive neuron loss in adult mice. Hq mutant mice are characterized initially by a loss of hair in homozygous females and hemizygous males. These mice develop progressive ataxia concomitant with loss of cerebellar neurons. Cell loss in Hq mutants peaks at 5-7 months and is initially confined to granule cells in the caudal cerebellum. Analysis of cell cycle markers demonstrates that granule cell apoptosis is accompanied by abortive cell cycle re-entry. These abnormally cycling cells express sonic hedgehog, a potent mitogen expressed by granule cell precursors, but down-regulated upon terminal differentiation of these cells. The Hq critical region has been refined to a 0.61 cM region of the X Chromosome, and a genomic contig across this region has been assembled. Genetic analysis of transcripts within the Hq region demonstrated that the gene encoding the apoptosis-inducing factor (Aif ), a mitochondrial oxidoreductase, cosegregates with the Hq gene. Further, Aif transcript levels are greatly reduced in pre-ataxic mutant mice, making Aif a likely candidate for the Hq gene. Immunohistochemistry results demonstrate oxidized DNA is present in mutant but not control granule cells, suggesting down-regulation of Aif results in oxidative stress. Experiments outlined in this grant will identify the molecular lesion in the Aif gene in Hq mutant mice. In addition, the effect of the Hq mutation on mitochondrial function and oxidative stress will be analyzed. Lastly, to test whether the ectopic expression of sonic hedgehog is sufficient to cause cell cycle re-entry and subsequent apoptosis, transgenic mice will be generated that abnormally express this molecule in terminally differentiated granule cells. The results of these experiments will allow validation of Hq mutant mice as a model for elucidation of the molecular interplay between oxidative stress, mitogen activation and cell cycle re-entry, and neuronal death in the adult nervous system.